Tyrosine kinase receptors play a central role in oncogenesis and other pathophysiologic processes as well as in normal growth and development. When activated by ligand these receptors induce a diverse array of responses, and these responses differ between cell types expressing the same receptor and between different receptor types expressed in the same cell. The important roles of autophosphorylation and interaction with SH2 domains in signalling by tyrosine kinase receptors are well established. However, it is uncertain whether this accounts for all of the pleiotropic effects of these receptors. In this application we propose an examination of two alternative pathways for EGF receptor activation and signalling. I. Phosphorylation and activation of the EGF receptor by pp60src. We have found that the EGF receptor in src transformed cells is constitutively phosphorylated on novel sites and partially activated. The goals to extend this analysis are: A. Identification of the novel sites of src-induced phosphorylation. B. Analysis of the function of those phosphorylation sites in EGF receptor signalling. C. Determination of the role of these phosphorylation sites in the synergistic biological interactions between pp60c-src and the EGF receptor. II. Signalling by kinase-defective EGF receptors. Although it has generally been thought that tyrosine kinase receptors depend on their kinase activity for cell signalling, this may not be true in all cases: it seems clear that a kinase-defective EGF receptor can activate MAP kinase. This activity may represent a hitherto undiscovered signalling capacity of this tyrosine kinase receptor (and perhaps other tyrosine kinase receptors) which has been undetected because of the prominence of the kinase-dependent signalling systems. We propose to investigate the biochemical mechanisms by which the EGF receptor activates MAP kinase, moving "downstream" from the receptor and "upstream" from MAP kinase: A. Analysis of signalling proteins which may interact directly with the kinase-defective EGF receptor, including tyrosine kinases, p21ras and heterotrimeric G-proteins. B. Investigation of the specific upstream components leading to activation of MAP kinase by kinase-defective receptors, including MAP Kinase Kinases (MEKs), raf and STE11 (MEK kinases).